The RCM technique extends standard confocal microscopy with a re-scanning unit, improving lateral resolution by √2 and increasing the signal to noise ratio.
Re-scan Confocal Microscopy (RCM) is a new super-resolution technique based on standard confocal microscopy extended with an optical (re-scanning) unit that projects the image directly on a CCD- or sCMOS camera. This new microscope has improved lateral resolution (170 nm at 488 nm excitation), and strongly improved sensitivity, while maintaining the sectioning capability of a standard confocal microscope. It is particularly useful for biological applications where the combination of high-resolution and high-sensitivity is required.
During scanning, re-scan mirrors (SM2) have a bigger amplitude than the first scan mirrors (SM1). This magnifies the image on the camera chip compared to the sample, and eventually results in the higher resolution of the image. The resolution of the system is improved with the re-scan step by a factor of √2 (i.e. 1.41 times), compared to Abbe’s resolution limit by changing the angular amplitude of the re-scanner (SM2). Closing down the pinhole is no longer necessary to increase resolution. Since the re-scan is a purely optical method with no further image processing required, there is no cost in time while improving the resolution. By using a sensitive camera as detector, the signal-to-noise ratio of the RCM is 4 times higher than in standard PMT-based confocal microscope.
To fully understand the principle of rescanning, resolution improvement and the optical layout of the RCM, please watch the video below that explains the components and the light path of the RCM (animation credits to StudioFlip). Additional technical details and test images can be found in De Luca et al (2013).